मानव आंत्र mucosa Explants की संस्कृति और फूट डालना उत्तेजना के लिए एक उपन्यास विधि
Summary
हम कम से कम 24 घंटे से अधिक उत्तेजनाओं की विभिन्न प्रकार की प्रतिक्रिया की संस्कृति और निगरानी में मानव आंत्र mucosa के रखरखाव के लिए एक उपन्यास विधि परिचय. हमारे विधि के साथ, ऊतक के polarity शिखर मार्ग के माध्यम से एक शारीरिक उत्तेजना के लिए अनुमति दी जाती है.
Abstract
Few models currently exist to realistically simulate the complex human intestine’s micro-environment, where a variety of interactions take place. Proper homeostasis directly depends on these interactions, as they shape an entire immunological response inducing tolerance against food antigens while at the same time mounting effective immune responses against pathogenic microbes accidentally ingested with food.
Intestinal homeostasis is preserved also through various complex interactions between the microbiota (including food-associated beneficial bacterial strains) and the host, that regulate the attachment/degradation of mucus, the production of antimicrobial peptides by the epithelial barrier, and the “education” of epithelial cells’ that controls the tolerogenic or immunogenic phenotype of unique, gut-resident lymphoid cells’ populations. These interactions have been so far very difficult to reproduce with in vitro assays using either cultured cell lines or peripheral blood mononuclear cells. In addition, mouse models differ substantially in components of the intestinal mucosa (mucus layer organization, commensal bacteria community) with respect to the human gut. Thus, studies of a variety of treatments to be brought in the clinics for important stress-related or pathological conditions such as irritable bowel syndrome, inflammatory bowel disease or colorectal cancer have been difficult to carry out.
To address these issues, we developed a novel system that enables us to stimulate explants of human intestinal mucosa that retain their in situ conditioning by the host microbiota and immune response, in a polarized fashion. Polarized apical stimulation is of great importance for the outcome of the elicited immune response. It has been repeatedly shown that the same stimuli can produce completely different responses when they bypass the apical face of the intestinal epithelium, stimulating epithelial cells basolaterally or coming into direct contact with lamina propria components, switching the phenotype from tolerogenic to immunogenic and causing unnecessary and excessive inflammation in the area.
We achieved polarized stimulation by gluing a cave cylinder which delimited the area of stimulation on the apical face of the mucosa as will be described in the protocol. We used this model to examine, among others, differential effects of three different Lactobacilli strains. We show that this model system is very powerful to assess the immunomodulatory properties of probiotics in healthy and disease conditions.
Protocol
Representative Results
Discussion
मानव आंत्र mucosa के लिए उपचार सप्रमाण परीक्षण किया जा सकता है जिस पर physiologically प्रासंगिक मॉडल के लिए की जरूरत लंबे समय से जोर दिया गया है. कई शोधकर्ताओं ने उस प्रयोजन के लिए अब तक का इस्तेमाल सेल 5 और माउस मॉ?…
Declarações
The authors have nothing to disclose.
Acknowledgements
हम ऑक्सीजन कक्षों प्रदान करने के लिए उत्कृष्ट तकनीकी सहायता और डॉ. एंटोनियो डि Sabatino के लिए एरिका Mileti धन्यवाद.
धन: एमआर को और Fondazione Cariplo और एक मैरी क्यूरी अंतरराष्ट्रीय प्रशिक्षण गतिशीलता नेटवर्क (क्रॉस टॉक, अनुदान समझौता नहीं: 21553-2) द्वारा समर्थन करने के लिए: इस काम के 7 वें यूरोपीय संघ के ढांचे कार्यक्रम (Dendroworld IBDase, ईआरसी) से अनुदान द्वारा समर्थित किया गया के.टी..
Materials
| Regeant | Company | Catalog No. | |
| Reagents | |||
| 10X HBSS | Eurocalne | ECM4006XL | |
| Peni/Strepto | Lonza | DE17602E | |
| Gentamycin | Gibco | 15750-037 | |
| DMEM | Lonza | BE12614 | |
| FBS-Na | Gibco | 16000-044 | |
| 100X ITS-X | Gibco | 51500-056 | |
| EGF | Tebu-bio | 100-15 | |
| L-Glutamine | Lonza | BE17605E | |
| Non essential aminoacids 100X | Gibco | 11140-035 | |
| NaPyr | Gibco | 11360-039 | |
| Materials | |||
| Cloning cylinders 6×8 mm | BellCo | 2090-00608 | |
| Cloning cylinders 8×8 mm | BellCo | 2090-0080 | |
| Cloning cylinders 10×10 mm | BellCo | 2090-01010 | |
| Metal grids | Home made | ||
| Centre well organ culture plate | BD Falcon | 353037 | |
| Surgical glue (Vetbond) | 3M | 1469SB | |
| Oxygen Jars | Home made | ||
| Oxygen tanks | Air Liquid Sanità |
Referências
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